The goal of this research effort is to understand how an important type of white blood cell, called a T lymphocyte, recognizes the presence of a microorganism or cancer cell in the body, or inappropriately recognizes a normal component of the body (an "autoantigen"). This research examines this question at the protein and cell level, to provide a detailed understanding of how the substances (antigens) making up these microorganisms, cancer cells, or normal self-components, are made visible to the defending T cells or the autoreactive T cells that cause disease. To accomplish these goals, we have examined the structure and function of special cellular proteins called major histocompatibility complex (MHC) molecules that are essential for antigens to be recognized by T cells. Specific questions involve understanding how structural differences in the MHC molecules of different individuals affect the ability of these proteins to stimulate T cells, the exact nature of the antigens bound to the MHC molecules, and the events within a cell that bring the antigen and MHC molecule together to form the complex needed to activate a T cell. These events are termed antigen processing and presentation. By understanding these events, we will be able to identify the most important components of microorganisms or cancer cells to use in protective or therapeutic vaccines, the best form in which to expose the body to these substances to stimulate an effective immune response, and possible ways in which to interfere with this recognition system, so that we can interrupt unwanted responses to "self" that cause autoimmune diseases such as multiple sclerosis, diabetes, or rheumatoid arthritis. Our studies have helped show that the two different classes of MHC proteins are specialized for binding to these antigenic peptides at different places inside a cell. This specialization helps the immune system find antigens whether the invading organism lives inside a cell or outside it, or whether the tumor mutation is in an internal or external protein. Our current work has focused on the special properties of MHC class II molecules and of associated proteins that help them capture and present peptide antigens found in small vesicles inside the cell where certain bacteria such as M. tuberculosis often live. During this past year, we have uncovered the mechanism by which a special protein molecule termed the invariant chain helps class II MHC molecules assemble inside the cell and remain functional until they can capture antigenic peptides in these vesicles. We have also identified a new traffic-control signal in MHC class II molecules that regulates the movement of these molecules into the antigen-containing vesicles even when invariant chain isn't present. Our new work has revealed how these traffic-control signals in invariant chain and in class II molecules function together to help the immune system recognize a maximum number of peptides from each antigenic protein. Finally, we have produced a new type of tool for studying antigen presentation by MHC class I and class II molecules. Special monoclonal antibodies that specifically bind only to selected antigenic peptide - MHC molecule combinations have been produced and used to study how antigen presentation takes place inside living cells and even in intact animals. These new reagents will help explore aspects of antigen presentation in living animals that were previous inaccessible to experimentation.